Graduating senior Tin Vo, of Jersey City, received the second-place silver undergraduate medal for his project, “Altering Substrate Specificity of a-Ketoglutarate Dehydrogenase for Chemical Synthesis,” from the NJIT Dana Knox Student Research Showcase competition on April 14, 2010. Professor Edgardo Farinas, PhD, was his advisor.
The long-term goal was to synthesize thiol esters from simple starting materials such as racemic amino acids. The aim was to engineer the multi-enzyme complex [alphasymbol]-ketogluarate dehydogenase (OGDHc). OGDHc is part of the critic acid cycle, which is the final common pathway for the oxidation of fuel molecules that include carbohydrates, fatty acids and amino acids.
OGDHc consist of three components: E1o, E2o and E3. E1o is responsible for substrate recognition and decarboxylation of the natural substrate a-ketoglutarate. E1o was engineered to accept 2-oxovaleric acid, which substitutes a hydrophobic (-CH3) group for an acidic (-CO2-) substituent found in the natural substrate. His260 and His298 were identified to be key residues in recognition of the acidic functional group.
Libraries of His260, His298, His260/His298 were screened for altered substrate specificity. Screening the His298 library identified seven possible mutants with activity towards 2-oxovaleric acid. However, the His260 library did not yield any positive mutants. Currently, a rescreen is being performed to eliminate false positives.